The binding characteristics and utilization of Aleuria aurantia,Lens culinaris and few other lectins in the elucidation of fucosyltransferase activities resembling cloned FT VI and apparently unique to colon cancer cells

Human colon carcinoma cell fucosyltransferase (FT) in contrast to the FTs of several human cancer cell lines, utilized GlcNAcbeta1,4GlcNAcbeta-O-Bn as an acceptor, the product being resistant to alpha1,6-L-Fucosidase and its formation being completely inhibited by LacNAc Type 2 acceptors. Further, this enzyme was twofold active towards the asialo agalacto glycopeptide as compared to the parent asialoglycopeptide. Only 60% of the GlcNAc moieties were released from [14C]fucosylated asialo agalacto triantennary glycopeptide by jack bean beta-N-acetylhexosaminidase. These alpha1,3-L-fucosylating activities on multiterminal GlcNAc residues and chitobiose were further examined by characterizing the products arising from fetuin triantennary and bovine IgG diantennary glycopeptides and their exoglycosidase-modified derivatives using lectin affinity chromatography. Utilization of [14C]fucosylated glycopeptides with cloned FTs indicated that Lens culinaris lectin and Aleuria aurantia lectin (AAL) required, respectively, the diantennary backbone and the chitobiose core alpha1,6-fucosyl residue for binding. The outer core alpha1,3- but not the alpha-1,2-fucosyl residues decreased the binding affinity of AAL. The AAL-binding fraction from [14C]fucosylated asialo fetuin, using colon carcinoma cell extract, contained 60% Endo F/PNGaseF resistant chains. Similarly AAL-binding species from [14C]fucosylated TFA-treated bovine IgG using colon carcinoma cell extract showed significant resistance to endo F/PNGaseF. However, no such resistance was found with the corresponding AAL non- and weak-binding species. Thus colon carcinoma cells have the capacity to fucosylate the chitobiose core in glycoproteins, and this alpha1,3-L-fucosylation is apparently responsible for the AAL binding of glycoproteins. A cloned FT VI was found to be very similar to this enzyme in acceptor substrate specificities. The colon cancer cell FT thus exhibits four catalytic roles, i.e., alpha1,3-L-fucosylation of: (a) Galbeta1,4GlcNAcbeta-; (b) multiterminal GlcNAc units in complex type chain; (c) the inner core chitobiose of glycopeptides and glycoproteins; and (d) the nonreducing terminal chiotobiose unit.     

A sialic acid-specific lectin from the slug Limax flavus

The slug, Limax flavus, contains a lectin that appears to be highly specific for sialic acid residues of glycoproteins. The carbohydrates which inhibited the hemagglutinating activity of the slug lectin and the concentration of the carbohydrate which gave a 50% inhibition are as follows: N-acetylneuraminic acid, 0.13 mm; N-glycolylneuraminic acid, 0.90 mm; d-glucosamine, 4.9 mm; d-galactosamine, 7.6 mm; N-acetyl-d-glucosamine, 23 mm; and N-acetyl-d-galactosamine, 24 mm. d-Galactose, d-glucose, d-mannose, α-methyl-d-glucoside, α-methyl-d-mannoside, l-arabinose, d-xylose, l-fucose, d-glucuronic acid, lactose, and sucrose were found to be ineffective as inhibitors of the hemagglutinating activity of the slug lectin. Hemagglutination by slug lectin was strongly inhibited by bovine submaxillary mucin and fetuin but not by sialic acid-free bovine submaxillary mucin or fetuin.     

Mucin binding mitogenic lectin from freshwater Indian gastropod Belamyia bengalensis: purification and molecular characterization

A lectin was purified from the hemolymph of the freshwater Indian gastropod Belamyia bengalensis. The purification involved successive ion-exchange chromatography on Resource Q and gel filtration on Superose 12 column in FPLC system. Homogeneity of the protein was confirmed by polyacrylamide gel electrophoresis. Belamyia bengalensis lectin (BBL) was a monomeric protein with a molecular weight of 33 kDa as demonstrated by gel filtration and SDS-PAGE. It is a glycoprotein containing 6% total sugar and its activity is highly dependent on Ca(2+). BBL agglutinated human erythrocytes and is a blood group non-specific lectin. It agglutinated animal erythrocytes also. Hapten inhibition studies indicated that BBL shows binding specificity only for N-acetyl-D-glucosamine and N-acetyl-D-galactosamine at a high concentration among the mono- and oligosaccharides tested. Among the glycoproteins used for hemagglutination-inhibition assay, porcine submaxillary mucin was found to be the best inhibitor. Chemical modification studies indicated that Lys, Arg, and Trp are essential for the sugar-binding activity of BBL. Circular dichroism spectra revealed high content of alpha-helical structure in the lectin. BBL is a potent mitogen as it stimulated the T-lymphocyte proliferation, specifically the Th1 subset.     

Sialic acid is an essential moiety of mucin as a hydroxyl radical scavenger

In this work, we examined the antioxidant role of mucin, a typical sialic acid containing high-molecular weight glycoprotein. The function of mucin as a hydroxyl radical (.OH) scavenger was characterized using bovine submaxillary gland mucin (BSM). Non-treated BSM effectively protected DNA from the attack of .OH; however, desialylated BSM lost this potential. Moreover, we estimated the scavenging effects of BSM against .OH generated by UV irradiation of hydrogen peroxide using ESR analysis. Our results indicate that BSM has .OH scavenging ability the and sialic acid in mucin is an essential moiety to scavenge .OH.     

Purification and characterization of a sialic acid specific lectin from the hemolymph of the freshwater crab Paratelphusa jacquemontii.

A naturally occurring hemagglutinin was detected in the serum of the freshwater crab, Paratelphusa jacquemontii (Rathbun). Hemagglutination activity with different mammalian erythrocytes suggested a strong affinity of the serum agglutinin for horse and rabbit erythrocytes. The most potent inhibitor of hemagglutination proved to be bovine submaxillary mucin. The lectin was purified by affinity chromatography using bovine submaxillary mucin-coupled agarose. The molecular mass of the purified lectin was 34 kDa as determined by SDS/PAGE. The hemagglutination of purified lectin was inhibited by N-acetylneuraminic acid but not by N-glycolylneuraminic acid, even at a concentration of 100 mm. Bovine submaxillary mucin, which contains mainly 9-O-acetyl- and 8,9 di-O-acety-N-acetyl neuraminic acid was the most potent inhibitor of the lectin. Sialidase treatment and de-O-acetylation of bovine submaxillary mucin abolished its inhibitory capacity completely. Also, asialo-rabbit erythrocytes lost there binding specificity towards the lectin. The findings indicated an O-acetyl neuraminic acid specificity of the lectin.     

Structural basis for the energetics of jacalin-sugar interactions: promiscuity versus specificity

Jacalin, a tetrameric lectin, is one of the two lectins present in jackfruit (Artocarpus integrifolia) seeds. Its crystal structure revealed, for the first time, the occurrence of the beta-prism I fold in lectins. The structure led to the elucidation of the crucial role of a new N terminus generated by post-translational proteolysis for the lectin's specificity for galactose. Subsequent X-ray studies on other carbohydrate complexes showed that the extended binding site of jacalin consisted of, in addition to the primary binding site, a hydrophobic secondary site A composed of aromatic residues and a secondary site B involved mainly in water-bridges. A recent investigation involving surface plasmon resonance and the X-ray analysis of a methyl-alpha-mannose complex, had led to a suggestion of promiscuity in the lectin's sugar specificity. To explore this suggestion further, detailed isothermal titration calorimetric studies on the interaction of galactose (Gal), mannose (Man), glucose (Glc), Me-alpha-Gal, Me-alpha-Man, Me-alpha-Glc and other mono- and oligosaccharides of biological relevance and crystallographic studies on the jacalin-Me-alpha-Glc complex and a new form of the jacalin-Me-alpha-Man complex, have been carried out. The binding affinity of Me-alpha-Man is 20 times weaker than that of Me-alpha-Gal. The corresponding number is 27, when the binding affinities of Gal and Me-alpha-Gal, and those of Man and Me-alpha-Man are compared. Glucose (Glc) shows no measurable binding, while the binding affinity of Me-alpha-Glc is slightly less than that of Me-alpha-Man. The available crystal structures of jacalin-sugar complexes provide a convincing explanation for the energetics of binding in terms of interactions at the primary binding site and secondary site A. The other sugars used in calorimetric studies show no detectable binding to jacalin. These results and other available evidence suggest that jacalin is specific to O-glycans and its affinity to N-glycans is extremely weak or non-existent and therefore of limited value in processes involving biological recognition.     

Specificity of the sialic acid-binding lectin from the snail Cepaea hortensis.

The specificity of the sialic acid-binding lectin from the snail Cepaea hortensis, purified by affinity chromatography on fetuin-Sepharose, was studied by hemagglutination inhibition assay applying 32 sialic acid derivatives and 14 glycoproteins. 2-alpha-Methyl-9-O-acetyl-NeuAc was the most potent inhibitor, followed closely by 2-alpha-methyl-NeuAc and 2-alpha-benzyl-NeuAc. An axially orientated carboxyl group is a prerequisite for maximal lectin-sugar binding. Neither size nor polarity of the alpha-anomeric substituent significantly influenced inhibition potency. An intact sialic acid N-acetyl group is essential for optimal lectin-sugar interaction. The trihydroxypropyl side chain also is of great importance. However, a bulky hydrophobic substituent at the side chain like a 9-O-tosyl residue did not decrease binding to the lectin. The lectin did not distinguish between NeuAc alpha 2----3Gal beta 1----4Glc and NeuAc alpha 2----6Gal beta 1----4Glc. Among other sugars tested, only N-acetylglucosamine showed inhibition, although 50-fold less. The most potent glycoprotein inhibitors were those carrying O-chains only or preferentially, as ovine submaxillary mucin, bovine submaxillary mucin, and glycophorin A. Tamm-Horsfall protein was an exception being a strong inhibitor, although carrying only N-chains. Asialoglycoproteins were inactive. Glycoproteins containing the NeuAc alpha 2----3Gal sequence inhibited the lectin as well as those with NeuAc alpha 2----6GalNAc. From the results a model of the lectin's binding site for sialic acid is suggested.     

Specificity of submaxillary gland sialyltransferases

A method has been developed to determine the activities of specific sialyltransferases by analysis of the products of the reaction. This method, which utilizes high performance liquid chromatography, distinguishes addition of sialic acid to the N-acetylgalactosamine vs. galactose residues of the mucin disaccharide Galβ(1→3)GalNac, and can be used to distinguish formation of the 3′- and 6′-isomers of sialyllactose. For the bovine, ovine, and porcine submaxillary extracts, more than 95% of the activity with asialo ovine submaxillary mucin is due to formation of NeuAc α(2→6)GalNAc. With lactose as the acceptor, more than 95% of the α(2→3) isomer is produced. Activity with asialofetuin is due solely to the O-linked chain, with relative activity toward the galactose vs. GalNAc residues of 0.32, 1.5, and 0.10 for bovine, ovine, and porcine, respectively. The rat submaxillary gland extract showed equal formation of 3′- and 6′-sialyllactose, and very low activity with asialo ovine submaxillary mucin. However, at least 40% of the activity toward the Galβ(1→3)GalNAc disaccharide of asialofetuin was directed toward the GalNAc residue. The relative preference of the N-acetylgalactosaminide α(2→6) sialyltransferase for a monosaccharide vs. a substituted GalNAc may play a role in regulation of chain length during mucin synthesis.     

Multivalent II [beta-D-Galp-(1-->4)-beta-D-GlcpNAc] and Talpha [beta-D-Galp-(1-->3)-alpha-D-GalpNAc] specific Moraceae family plant lectin from the seeds of Ficus bengalensis fruits

A galactose specific lectin was isolated from the seeds of Ficus bengalensis (Moraceae) fruits and designated as F. bengalensis agglutinin (FBA). The lectin was purified by affinity repulsion chromatography on fetuin-agarose and was a monomer of molecular mass 33kDa. Like other Moraceae family lectins, carbohydrate-binding activity of FBA was independent of any divalent cation. FBA did not bind with simple saccharides, however sugar ligands with aromatic aglycons showed pronounced binding. The combining site of FBA recognized preferably Galbeta1,4GlcNAcbeta1-(II) followed by Galbeta1,3GalNAcalpha1-(Talpha) containing glycotopes. Interaction with saccharides revealed that the combining site of FBA could well accommodate a tetrasaccharide, asialo GM1 glycan (Galbeta1,3GalNAcbeta1,4Galbeta1,4Glcbeta1-), whereas polyvalent Tn (GalNAcalpha1-Ser/Thr), one of the well-recognized ligands of Moraceae family lectin, did not interact well with FBA.     

Purification and characterization of lectin from fruiting body of Ganoderma lucidum: Lectin from Ganoderma lucidum

A novel 114 kDa hexameric lectin was purified from the fruiting bodies of the mushroom Ganoderma lucidum. Biochemical characterization revealed it to be a glycoprotein having 9.3% neutral sugar and it showed hemagglutinating activity on pronase treated human erythrocytes. The lectin was stable in the pH range of 5–9 and temperature up to 50 °C. The hemagglutinating activity was inhibited by glycoproteins that possessed N-as well as O-linked glycans. Chemical modification of the G. lucidum lectin revealed contribution of tryptophan and lysine to binding activity. The thermodynamics of binding of bi- and triantennary N-glycans to G. lucidum lectin was studied by spectrofluorimetry. The lectin showed very high affinity for asialo N-linked triantenary glycan and a preference for asialo glycans over sialylated glycans. The binding was accompanied with a large negative change in enthalpy as well as entropy, indicating primarily involvement of polar hydrogen, van der Waals and hydrophobic interactions in the binding.     

Single‐molecule pair studies of the interactions of the α‐GalNAc (Tn‐antigen) form of porcine submaxillary mucin with soybean agglutinin

Mucins form a group of heavily O‐glycosylated biologically important glycoproteins that are involved in a variety of biological functions, including modulating immune response, inflammation, and adhesion. Mucins are also involved in cancer and metastasis and often express diagnostic cancer antigens. Recently, a modified porcine submaxillary mucin (Tn‐PSM) containing GalNAcα1‐O‐Ser/Thr residues was shown to bind to soybean agglutinin (SBA) with ～10⁶‐fold enhanced affinity relative to GalNAcα1‐O‐Ser, the pancarcinoma carbohydrate antigen. In this study, dynamic force spectroscopy is used to investigate molecular pairs of SBA and Tn‐PSM. A number of force jumps that demonstrate unbinding or rebinding events were observed up to a distance equal to 2.0 μm, consistent with the length of the mucin chain. The unbinding force increased from 103 to 402 pN with increasing force loading rate. The position of the activation barrier in the energy landscape of the interaction was 0.1 nm. The lifetime of the SBA–TnPSM complex in the absence of applied force was determined to be in the range 1.3–1.9 s. Kinetic parameters describing the rate of dissociation of other sugar lectin interactions are in the range 3.3 × 10^?3–2.5 × 10^?3 s. The long lifetime of the SBA‐TnPSM complex is compatible with a binding model in which lectin molecules “bind and jump” from α‐GalNAc residue to α‐GalNAc residue along the polypeptide chain of Tn‐PSM before dissociating. These findings have important implications for the molecular recognition properties of mucins. © 2009 Wiley Periodicals, Inc. Biopolymers 91: 719–728, 2009. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com     

Purification and characterization of a lectin from endophytic fungus Fusarium solani having complex sugar specificity

A lectin from the mycelial extract of an endophytic strain of Fusarium solani was purified. Its hemagglutinating activity was inhibited by glycoproteins possessing N-linked as well as O-linked glycans. The thermodynamics and kinetics of binding of glycans and glycoproteins to F. solani lectin was studied using surface plasmon resonance. The lectin showed high affinity for asialofetuin, asialomucin, asialofibrinogen, and thyroglobulin; and comparatively low affinity for mucin, fetuin, fibrinogen, and holotransferrin. Glycoproteins showed several fold higher affinity than their corresponding glycans with significant contribution from enthalpy and positive entropy, suggesting the involvement of non-polar protein-protein interaction. Moreover, the higher affinity of the glycoproteins was due to their faster association rates and low activation energy.     

Characterization of a new α-galactosyl-binding lectin from the mushroom Clavaria purpurea

A galactose specific lectin (CpL) was purified from the Clavaria purpurea mushroom by affinity chromatography. CpL agglutinated only trypsin-treated rabbit erythrocytes. On enzyme linked lectin sorbent assay (ELLSA), the lectin bound with thyroglobulin and asialo bovine submaxillary mucin (BSM). The fine sugar binding specificity of CpL was elucidated using inhibition of hemagglutination and sugar immobilized gold nano-particles (SGNP). The results indicated a preference of CpL towards α-galactosyl sugar chains. Among several monosaccharides and disaccharides assayed for dissociation effect on the SGNP-CpL complex, Galα1-3Gal and raffinose were the best inhibitors. The partial amino acid sequence showed two QXW motifs in CpL and similarity towards members of the ricin B superfamily.     

Purification and characterization of an N-acetyl-d-galactosamine-specific lectin from the edible mushroom Schizophyllum commune

An N-acetyl-d-galactosamine (GalNAc)-specific lectin was purified from the edible mushroom, Schizophyllum commune, using affinity chromatography on a porcine stomach mucin (PSM)-Sepharose 4B column. Under reducing and non-reducing conditions, SDS-polyacrylamide gel electrophoresis gave a major band of 31.5 kDa. The Schizophyllum commune lectin (SCL) showed high affinity toward rat erythrocytes and the sugar inhibition assay exhibited its sugar specificity highly toward lactose and N-acetyl-d-galactosamine. It was stable at 55 °C for 30 min and at pH 3–10 for 18-h test. The lectin was shown to be a glycoprotein with cytotoxic activity against human epidermoid carcinoma cells. The N-terminus of SCL was blocked but amino acid sequences of internal tryptic peptides showed moderately sequence similarities with some other fungal and plant lectins. Crystals of SCL were obtained by the sitting drop vapour-diffusion method using polyethylene glycol 8000 as the precipitant, and gave an X-ray diffraction pattern to approximately 3.8 Å resolution.     

A sialic acid-binding lectin from ovine placenta: purification,specificity and interaction with actin

A sialic-acid-specific lectin from ovine placental cotyledons was purified by affinity chromatography on bovine submaxillary mucin-agarose followed by gel filtration, and it showed a molecular weight of 65 000 by sodium dodecylsulfate-polyacrylamide gel electrophoresis. This lectin has the capacity to interact with actin, since in binds to actin-F in a cosedimentation assay and it acts as a mediator in the binding of action to the affinity column. The lectin agglutinated rabbit and rat erythrocytes, but not human A, B or O erythrocytes. Haemagglutination inhibition assays of different saccharides, glycoproteins and glycolipids indicate that this lectin has affinity for sialic acid, which is enhanced by its O-acetylation. The N-terminal sequence of the protein shows 92% identity with rabbit and porcine uterine calreticulin.     

Analysis of the carbohydrate binding specificity of the mushroom Pleurotus ostreatus lectin by surface plasmon resonance

The sugar binding specificity of the mushroom Pleurotus ostreatus lectin (POL) was analyzed by surface plasmon resonance. The lectin was immobilized to a sensor chip, and asialo-bovine submaxillary mucin (asialo-BSM), one of the most potent inhibitors in the hemagglutination inhibition assay, tightly bound to the lectin. The binding specificity of various mono- or oligosaccharides to the lectin was evaluated by the coinjection method. The dissociation of asialo-BSM was promoted by injection of some haptenic saccharides. For the most part, the order of acceleration ability of the sugars to the dissociation in the coinjection experiment agreed with that of the inhibitory potency of each sugar evaluated by the hemagglutination inhibition assay. In conclusion, POL recognized a galactosyl residue, and the specificity was increased by substitution at the C-2 position of the galactosyl residue with a fucosyl or acetylamino group. This method using the coinjection method proved useful in analysis of carbohydrate-lectin binding specificity.     

Recognition profile of Bauhinia purpurea agglutinin (BPA)

Bauhinia purpurea agglutinin (BPA) is a Galbeta1-3GalNAc (T) specific leguminous lectin that has been widely used in multifarious cytochemical and immunological studies of cells and tissues under pathological or malignant conditions. Despite these diverse applications, knowledge of its carbohydrate specificity was mainly limited to molecular or submolecular T disaccharides. Thus, the requirement of high density polyvalent or multi-antennary carbohydrate structural units for BPA binding and an updated affinity profile were further evaluated by enzyme-linked lectinosorbent (ELLSA) and inhibition assays. Among the glycoproteins (gps) tested and expressed as 50% nanogram inhibition, the high density polyvalent GalNAcalpha1-Ser/Thr (Tn) and Galbeta1-3/4GlcNAc (I/II) glycotopes present on macromolecules generated a great enhancement of binding affinity for BPA as compared to their monomers. The most potent inhibitors were a Tn-containing gp (asialo OSM) and a I/II containing gp (human blood group precursor gp), which were up to 1.7 x 10(4) and 2.3 x 10(3) times more potent than monovalent Gal and GalNAc, respectively. However, multi-antennary glycopeptides, such as tri-antennary Galbeta1-4GlcNAc, which was slightly more active than II or Gal, gave only a minor contribution. Regarding the carbohydrate structural units studied by the inhibition assay, blood group GalNAcbeta1-3/4Gal (P/S) active glycotopes were active ligands. The overall binding profile of BPA was: high density polyvalent T/Tn and II clusters > Tn-glycopeptides (M.W. <3.0 x 10(3))/Talpha monomer > monovalent P/S > Tn monomer and GalNAc > tri-antennary II > Gal > Man and Glc (inactive). These findings give evidence for the binding of this lectin to dense cell surface T, Tn and I/II glycoconjugates and should facilitate future usage of this lectin in biotechnological and medical applications.     

Lectins in the hemolymph of Japanese horseshoe crab, Tachypleus tridentatus

The hemolymph of the Japanese horsehoe crab, Tachypleus tridentatus contains lectins which agglutinate mammalian erythrocytes. Affinity chromatographic purification of the lectins using bovine submaxillary gland mucin-conjugated Sepharose resulted in the separation of the lectins into four fractions; one major and three minor lectins. Protein subunits revealed by polyacrylamide gel electrophoresis and the immunoprecipitin line of these lectins against antiserum to crude lectins were unique to each fraction. The activities of all the lectins were optimal at pH values between 6 and 8, and were destroyed by heating at 60°C. Calcium chloride augumented the activities of three lectins, but the major lectin was not influenced by the salt. Bovine erythrocytes were not agglutinated by any of the lectins and comparative agglutination titers for other erythrocytes from various sources were different among these lectins. The activities of all the lectins were inhibited by N-acetylamino sugars. They were more effectively inhibited by glycoproteins which contain sialic acid.     

A lectin from the Asian horseshoe crabTachypleus tridentatus: purification,specificity and interaction with tumour cells

A lectin from the haemolymph of the Asian horseshoe crabTachypleus tridentatus was purified to homogeneity by affinity chromatography on Sepharose 4B-boundN-acetylneuraminic acid. The specificity of this lectin was studied by haemagglutination inhibition with sialic acid analogues,N-acetylhexosamines and glycoproteins. For the interaction with the agglutinin theN-acetyl group and the glyceryl side chain ofN-acetylneuraminic acid are important, while presence of an aglycon, specially an -glycosidically linked lactose increases affinity to the lectin. The strongest glycoprotein inhibitors were ovine as well as bovine submaxillary mucin andCollocalia mucin, all beingO-chain glycoproteins but carrying completely different carbohydrate chains. The majority ofN-chain proteins were inactive. As the lectin agglutinates human erythrocytes, but not the murine lymphoma lines Eb and ESb or the human colon carcinoma HT 29, these cancer cells apparently lack the Tachypleus tridentatus agglutinin-receptor which is present on red cells andO-chain glycoproteins.Abbreviations TTA Tachypleus tridentatus agglutinin - SDS sodium dodecyl sulfate - BSM bovine sub-maxillary mucin - VCS Vibrio cholerae sialidase - OSM ovine submaxillary mucin - WGA Wheat germ agglutinin - NeuAc N-acetylneuraminic acid.     

Structural basis for the specificity of basic winged bean lectin for the Tn-antigen: a crystallographic, thermodynamic and modelling study

The crystal structure of winged bean basic agglutinin in complex with GalNAc-alpha-O-Ser (Tn-antigen) has been elucidated at 2.35 angstroms resolution in order to characterize the mode of binding of Tn-antigen with the lectin. The Gal moiety occupies the primary binding site and makes interactions similar to those found in other Gal/GalNAc specific legume lectins. The nitrogen and oxygen atoms of the acetamido group of the sugar make two hydrogen bonds with the protein atoms whereas its methyl group is stabilized by hydrophobic interactions. A water bridge formed between the terminal oxygen atoms of the serine residue of the Tn-antigen and the side chain oxygen atom of Asn128 of the lectin increase the affinity of the lectin for Tn-antigen compared to that for GalNAc. A comparison with the available structures reveals that while the interactions of the glyconic part of the antigen are conserved, the mode of stabilization of the serine residue differs and depends on the nature of the protein residues in its vicinity. The structure provides a qualitative explanation for the thermodynamic parameters of the complexation of the lectin with Tn-antigen. Modeling studies indicate the possibility of an additional hydrogen bond with the lectin when the antigen is part of a glycoprotein.